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  • Author or Editor: Robert E. Holland x
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Abstract

Objective—To examine the impact of simple versus complex extracellular matrices (ECMs) on morphologic development and differentiation of bovine mammary gland progenitor cells (BMGPCs).

Sample Population—Cultures of BMGPCs.

Procedures—BMGPCs were grown on the following extracellular matrices: collagen I, collagen IV, laminin, and a commercially available gelatinous protein mixture. Cells were examined with light microscopy and transmission electron microscopy.

Results—Formation of organoids and production of the gap junction protein, connexin 43, were the criteria for BMGPC differentiation. The BMGPCs formed a 2-dimensional monolayer when grown on plastic, laminin, collagen I, or collagen IV. These cells did not have a network of cells forming epithelial organoids resembling a honeycomb. However, they did produce gap junction proteins. When BMGPCs were cultured on the commercially available gelatinous protein mixture, 3-dimensional epithelial organoids resembling a honeycomb formed and connexin 43 was produced. The thickness of the commercially available gelatinous protein mixture also regulated cell shape reorganization. Cell density affected the formation organoid networks and the rate at which monolayers reached confluency.

Conclusions and Clinical Relevance—When plated on a commercially available gelatinous protein mixture, the BMGPC culture system allowed us to simulate, in vitro, the interaction between epithelial cells in varying stages of differentiation and the microenvironment. Thus, a heterogenous ECM, such as the commercially available gelatinous protein mixture, is more physiologically relevant in providing a microenvironment for BMGPC lineage pathway differentiation to mimic an in vivo environment. In contrast, BMGPCs grown on homogenous ECM, although able to produce connexin 43, are unable to form organoids.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To isolate bovine mammary gland cells with stem cell characteristics.

Sample Population—Monolayers of bovine mammary gland cells.

Procedure—Mammary gland cell populations were separated by use of selected media supplements. Phenotypic characteristics were examined via light and transmission electron microscopy. Cellular expression of casein and connexin 43 was identified immunohistochemically. A scrape-loading and dye transfer assay was used to examine the mammary gland cell populations for homogenous gap junctional intercellular communication (GJIC).

Results—Subpopulations of mammary gland cells grown in vitro are classified on the basis of their distinct morphologic features and ability to communicate via gap junctions. Ultrastructurally, 2 morphologically distinct cell types were classified as type I and II cells. Type I cells were small light undiffertiated cells and large light undifferentiated cells that were deficient in functional gap junctions (as is characteristic of stem cells). Type II cells included large light differentiated cells and terminally differentiated cells; GJIC was functional in type II cells. Type II cells had cytoplasmic expression of connexin 43, whereas, type I cells did not. All cells expressed casein.

Conclusions and Clinical Relevance—Subpopulations of bovine mammary gland cells with stem cell characteristics were identified. Phenotypic differences are observed among type I bovine mammary gland cells with stem cell characteristics. Gap junctional intercellular communication may be necessary for the differentiation of stem cells. Characterization of bovine mammary gland stem cells and their progeny may provide a new tool with which to study mammary gland health. (Am J Vet Res 2003;63:396–403)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine safety, efficacy, and immunogenicity of an intranasal cold-adapted modified- live equine influenza virus vaccine administered to ponies following induction of exercise-induced immunosuppression.

Design—Prospective study.

Animals—Fifteen 9- to 15-month old ponies that had not had influenza.

Procedure—Five ponies were vaccinated after 5 days of strenuous exercise on a high-speed treadmill, 5 were vaccinated without undergoing exercise, and 5 were not vaccinated or exercised and served as controls. Three months later, all ponies were challenged by nebulization of homologous equine influenza virus. Clinical and hematologic responses and viral shedding were monitored, and serum and nasal secretions were collected for determination of influenza-virus-specific antibody isotype responses.

Results—Exercise caused immunosuppression, as indicated by depression of lymphocyte proliferation in response to pokeweed mitogen. Vaccination did not result in adverse clinical effects, and none of the vaccinated ponies developed clinical signs of infection following challenge exposure. In contrast, challenge exposure caused marked clinical signs of respiratory tract disease in 4 control ponies. Vaccinated and control ponies shed virus after challenge exposure. Antibody responses to vaccination were restricted to serum IgGa and IgGb responses in both vaccination groups. After challenge exposure, ponies in all groups generated serum IgGa and IgGb and nasal IgA responses. Patterns of serum hemagglutination inhibition titers were similar to patterns of IgGa and IgGb responses.

Conclusions and Clinical Relevance—Results suggested that administration of this MLV vaccine to ponies with exercise-induced immunosuppression was safe and that administration of a single dose to ponies provided clinical protection 3 months later. (J Am Vet Med Assoc 2001;218:900–906)

Full access
in Journal of the American Veterinary Medical Association

Abstract

Objective—To evaluate serum haptoglobin concentration at feedlot arrival and subsequent performance and morbidity and mortality rates of calves that developed bovine respiratory disease.

Animals—360 heifer calves and 416 steer and bull calves.

Procedures—Serum samples were obtained from cattle at the time of arrival to a feedlot (day −1) and analyzed for haptoglobin concentration. In experiment 1, calves were classified into groups with a low (< 1.0 μg/mL), medium (1.0 to 3.0 μg/mL), or high (> 3.0 μg/mL) serum haptoglobin concentration and allotted into pens on the basis of group. In experiment 2, calves were classified as having or not having detectable serum haptoglobin concentrations.

Results—In experiment 1, average daily gain from days 1 to 7 decreased as haptoglobin concentration increased. Dry-matter intake (DMI) from days 1 to 21 decreased with increasing haptoglobin concentration, and DMI typically decreased from days 1 to 63. Total bovine respiratory disease morbidity rate typically increased with increasing haptoglobin concentration. At harvest, no differences in carcass characteristics were observed on the basis of haptoglobin concentration. In experiment 2, cattle with measureable serum haptoglobin concentrations at arrival weighed less throughout the experiment, gained less from days 1 to 7, and had lower DMI from days 1 to 42. Overall morbidity rate was not different between groups, but cattle with detectable serum haptoglobin concentrations had higher odds of being treated 3 times.

Conclusions and Clinical Relevance—Serum haptoglobin concentration in cattle at the time of feedlot arrival was not associated with overall performance but may have limited merit for making decisions regarding targeted prophylactic treatment.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine efficacy of a modified-live virus (MLV) vaccine containing bovine viral diarrhea virus (BVDV) 1a and 2a against fetal infection in heifers exposed to cattle persistently infected (PI) with BVDV subtype 1 b.

Animals—50 heifers and their fetuses.

Procedures—Susceptible heifers received a placebo vaccine administered IM or a vaccine containing MLV strains of BVDV1a and BVDV2a administered IM or SC. On day 124 (64 to 89 days of gestation), 50 pregnant heifers (20 vaccinated SC, 20 vaccinated IM, and 10 control heifers) were challenge exposed to 8 PI cattle. On days 207 to 209, fetuses were recovered from heifers and used for testing.

Results—2 control heifers aborted following challenge exposure; both fetuses were unavailable for testing. Eleven fetuses (8 control heifers and 1 IM and 2 SC vaccinates) were positive for BVDV via virus isolation (VI) and for BVDV antigen via immunohistochemical analysis in multiple tissues. Two additional fetuses from IM vaccinates were considered exposed to BVDV (one was seropositive for BVDV and the second was positive via VI in fetal tissues). A third fetus in the SC vaccinates was positive for BVDV via VI from serum alone. Vaccination against BVDV provided fetal protection in IM vaccinated (17/20) and SC vaccinated (17/20) heifers, but all control heifers (10/10) were considered infected.

Conclusions and Clinical Relevance—1 dose of a BVDV1a and 2a MLV vaccine administered SC or IM prior to breeding helped protect against fetal infection in pregnant heifers exposed to cattle PI with BVDV1b.

Full access
in American Journal of Veterinary Research